1. Field of the Invention
The present invention relates generally to the field of non-peptidyl inhibitors of protein tyrosine kinases. More particularly, the present invention concerns the inhibition of specific protein tyrosine kinases, for example pp60.sup.c-src, and growth factor receptors such as the fibroblast growth factor receptor. The instant inhibitors are based on a 1,4-benzodiazepin-2-one nucleus. The invention further relates to the use of these inhibitors in situations where the inhibition of a protein tyrosine kinase is indicated.
2. Description of Related Art
Protein tyrosine kinases (PTKs) are a family of enzymes which transfer the .gamma.-phosphate of ATP to the side chain of tyrosine residues on substrate proteins. There are three major classes of enzymes in this family. The first class to be identified were viral proteins, such as pp60.sup.v-src (the 60 kDa protein product of the viral src gene), which were capable of cell transformation. It was later discovered that these viral oncoproteins had normal cellular counterparts, for example pp60.sup.c-src (the 60 kDa protein product of the cellular src gene). These cellular proteins represent the second class of PTKs. The third class are represented by growth factor receptors, for example the fibroblast growth factor receptor (FGFr), the epidermal growth factor receptor (EGFr) and the platelet derived growth factor receptor (PDGFr).
PTKs are involved in a variety of different cellular processes. PTKs are often associated with cellular membranes and are involved in signal transduction and growth regulation. Alterations in the phosphorylation of PTK substrates are key events in cellular signaling.
Growth factor receptor tyrosine kinases are cell-surface proteins that receive information from outside the cell and convey it across the cell membrane. When a growth factor binds to its cognate receptor protein, the receptor is converted to an active form. The receptor then interacts with proteins on the inner surface of the cell membrane, altering their properties and/or subcellular location, resulting in changes in the behavior of the cell itself. These changes are many and various and may occur over the course of seconds (e.g., changes in ion flux), minutes (e.g., changes in cell shape), or hours (e.g., cellular mitosis).
Abberant levels, regulation and/or activity of a number of different PTKs play a key role in several human diseases. For example, pp60.sup.c-src (hereafter referred to as Src) is a PTK that is present in low levels, if at all, in normal eukaryotic cells (Barnekow, 1989; Punt et al., 1989). However, high concentrations and high activities of Src have been associated with numerous types of cancer. Breast cancer (Ottenhoff-Kalff et al., 1992; Partanen, 1994), stomach cancer (Takeshima et al., 1991), colon cancer (Rosen et al., 1986; Bolen et al., 1985, 1987; Cartwright et al., 1989, 1990; Talamonti et al., 1992, 1993; Termuhlen et al., 1993), hairy cell leukemia and a subgroup of B-cell lymphomas lymphomas (Lynch et al., 1993), low grade human bladder carcinoma (Fanning et al., 1992), and neuroblastoma (Bolen et al., 1985; O'Shaughnessy et al., 1987; Bjelfman et al., 1990) are all accompanied by greatly increased Src activity in the affected cells. Significantly, growth inhibition of several of these cancers has been correlated with decreases in Src activity, and for this reason, inhibitors of Src have excellent potential as anti-tumoral agents.
Other human cancers, e.g. melanoma, appear to be driven by an autocrine loop (Becker et al., 1989) comprising fibroblast growth factor (FGF) and its receptor (FGFr). As noted above, FGFr is a protein tyrosine kinase. An inhibitor of FGFr would therefore also be expected to have excellent potential as an anti-tumoral agent. Other serious diseases that have been linked with PTKs include osteoporosis, atherosclerosis, diabetic retinopathy, psoriasis, and possibly certain neurodegenerative disorders.
Inhibition of PTKs represents a promising avenue for treatment of disease. For example, Src is known to be involved with bone resorption (Hall et al., 1994), and it stands to reason that inhibition of Src would be an effective treatment modality in osteoporosis patents. Also, Src has recently been demonstrated to regulate the activity of N-methyl-D-aspartate (NMDA) ion channels (Yu et al., 1997) and NMDA-type glutamate channels in mammalian central neurons. Because dysfunction of NMDA receptor-mediated neurotransmission is believed to play a key role in the pathophysiology of Alzheimer's disease, Parkinson's disease (Coyle, et al. 1993), and schizophrenia (Heresco-Levy et al., 1996), this important discovery suggests that inhibition of Src may be helpful in the treatment of schizophrenia as well as other illnesses such as Parkinson's disease and seizure disorders that involve neural degeneration or abnormal regulation of NMDA channels.
There have been published reports of peptide-based inhibitors of PTKs. Various groups have substituted Phe or Phe analogues for Tyr in peptide substrates such as angiotensin II (Wong and Goldberg, 1984), epidermal growth factor receptor (EGFr) 1164-1176 (Honeggar et al., 1988), and insulin receptor (IR) 1142-1153 (Shoelson et al., 1989). Navarro et al. (1982) tested a series of peptidyl and amino acid halomethyl ketones as inhibitors of EGF-receptor kinase activity. Of nine compounds tested, only three showed significant inhibitory activity. Unfortunately, these investigators and others have found that peptides and oligonucleotides tend to have limited oral activities and rapid clearing times. Therefore, such materials have limited utility as bioavailable therapeutic agents. Thus, the development and characterization of non-peptidyl inhibitors of PTKs would represent a significant advance in the art.
Derivatives of 1,4-benzodiazepines have widespread biological activities and are one of the most important classes of bioavailable therapeutic agents (Sternbach, 1979). Benzodiazepine derivatives have been reported that are potent enzyme inhibitors, and that are agonists or antagonists, often with a high level of specificity and affinity. For example, 1,4-benzodiazepine derivatives have been described which are anxiolytic, anticonvulsant, and antihypnotic agents (Sternbach, 1979), selective cholecystokinin (CCK) receptor subtype A or B antagonists (Bock et al., 1989), K-selective opioid antagonists (Romer et al., 1982), platelet activating factor antagonists (Komecki et al., 1984), GP.sub.IIbIIIa inhibitors (Bondinell et al., 1993), and Ras farnesyl transferase inhibitors (James et al., 1993; Marsters et al., U.S. Pat. No. 5,532,359).
Unlike peptides, 1,4-benzodiazepines often have favorable pharmacokinetic properties such as good oral availability and long circulating half-lives. However, to date no suitable benzodiazepine inhibitors of PTKs have been described, and there are currently no specific potent small-molecule inhibitors of PTKs, such as Src, that have suitable pharmacokinetics, affinity, and specificity. Given the lack of inhibitors of protein tyrosine kinases with suitable properties, the development and characterization of non-peptidyl, small molecule inhibitors of PTKs would represent a significant contribution to the art.